Although more than 95% of dietary lipids are absorbed from the small intestine via the intestinal lymphatics, transported via the mesenteric collecting lymphatics through the lymphatic network en route to the blood, the role of lymph transport in the regulation of lipid metabolism is poorly understood. We have recently shown that high fructose diet-induced metabolic syndrome (MetSyn) rats exhibit impaired mesenteric lymphatic function. Dietary endotoxin readily associates with chylomicrons and are primarily absorbed through the mesenteric lymphatics along with many other gut peptides and hormones, suggesting that the primary trafficking of dietary endotoxins, such as, lipopolysaccharide (LPS) is via lymphatics. Hence, important questions remain unanswered: 1) what are the roles of lymphatic function in and responsiveness to dietary endotoxin and 2) how does that affect the progression of MetSyn. Our preliminary data show that immune cells, eosinophils and neutrophils are present on or adjacent to the lymphatic wall, and that the number of these immune cells on the lymphatic wall is decreased in LPS-induced inflamed mesenteric tissue. These data provide the basis for our central hypothesis: That dietary endotoxins alters the local cues (major cytokines of eosinophil and neutrophil: IL-4, IL-5, IL-8 and IL-13) in the mesenteric milieu that impairs lymphatic function and chylomicron transport, which leads to dyslipidemia and mesenteric adipose accumulation, both key pathologic determinants of the development of MetSyn. We will use the high-fructose and high-fat diet-induced MetSyn rat models and a LPS-induced inflammation model to determine the roles of the lymphatic system in the pathophysiology of MetSyn. Specific Aims are: 1) To demonstrate the linkages between dietary endotoxins, pro-inflammatory cytokines and lymphatic function during the development of MetSyn; 2) To determine the functional consequences and molecular mechanisms of reduced lymph transport in the development of MetSyn; and 3) To test the roles of micro RNAs (miRs) that interconnect the regulation of IL-8, IL-4, IL-5 and IL-13 in modulating lymphatic function during the development of MetSyn. We will use whole-mount mesenteric preparations for immunohistochemical studies, in situ preparations for lymph flow and lymphatic contractility, isolated/cannulated vessels for contractile characteristics of lymphatics, wire-myograph studies for length- tension and force-calcium measurements. These experiments will specifically address how dietary endotoxins effect the M1 and M2 macrophage polarization on lymphatics and the mechanisms by which the eosinophil and neutrophil interleukins, IL-4, IL-5, IL-8 and IL-13, and miR-19a, 93, 200c and 203 in lymphatic muscle cells regulate the lymphatic structure and function. Thus, this proposal will identify important targets bridging the mesentery milieu and lymphatic function, which could potentially provide diagnostic tools and/or therapies to improve lymph transport, and thus improve health in chronic inflammatory conditions such as MetSyn.